Studies on Drosophila innate immunity have highlighted the evolutionary conservation of response mechanisms with regards to bacterial and fungal infections. Indeed the role of the Toll receptors in mammalian immunity originated from studies on immunity in flies. In this proposal we aim to dissect the mechanisms of anti-viral innate immunity in Drosophila. In mammals, viruses trigger both acquired and innate immune responses. While the role of the innate immune system in anti-viral immunity is well established, many aspects of this response are still poorly understood. Because Drosophila has a mechanism to fight viruses, we propose to use this model organism, and take an unbiased genetic approach to identify the pathways that are required for viral immunity. Our hypothesis is that by studying immunity to viral pathogens in Drosophila, which does not have an acquired immune system, we will be able to identify conserved innate mechanisms by which organisms fight viruses. Viruses are a class of obligate intracellular pathogens that require host factors for their replication and survival. We have established both in vivo and in vitro assays to analyze the response of adult flies and cells to a Drosophila C picornavirus. We will use the powerful tools of genetics and functional genomics available in Drosophila to identify host factors involved in viral replication and immune protection to this RNA virus. Further, we will analyze the transcriptional response of host cells to viral infection to identify genes that are involved in the viral response. Using a robust tissue culture assay, which is based on the ability of the Drosophila C picornavirus to kill cells, we will conduct a highthroughput RNA interference screen to identify all proteins present in the Drosophila genome that when absent prevent viral infection. This assay will also be used to perform a chemical screen to identify small molecules that protect cells from viral infection. Because our insect virus system is sensitive to drugs that are used against human viruses, our studies may lead to the identification of novel antiviral compounds that may prove effective in mammalian cells.